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With the rapid rise in cases of COVID-19 across the world, health systems face unprecedented challenges in the delivery of patient care. This includes constrained capacity for intensive care unit (...

Background Inhaled antibiotics are standard of care for treating chronic pseudomonal respiratory infections in cystic fibrosis patients, initially approved for intermittent administration. However, use of continuous inhaled antibiotic regimens of differing combinations is growing. Methods This double-blind trial compared continuous alternating therapy (CAT) to an intermittent treatment regimen. Subjects were treated with 3cycles of 28-days inhaled aztreonam (AZLI) or placebo 3-times daily alternating with 28-days open-label tobramycin inhalation solution (TIS). Results 90 subjects were randomized over 18months. Study enrollment was limited, in part because of evolving practices by clinicians of adopting a CAT regimen in clinical practice; consequently the study was underpowered. AZLI/TIS treatment reduced exacerbation rates by 25.7% (p=0.25; primary endpoint) and rates of respiratory hospitalizations by 35.8% compared with placebo/TIS (p=0.14). AZLI/TIS CAT therapy was well tolerated. Conclusions This trial illustrates challenges with studying treatment regimens in a constantly evolving CF care environment. Nonetheless, the results of this trial indicate that AZLI/TIS CAT is well tolerated and may provide additional clinical benefit in CF patients compared with intermittent use of TIS alone. Clinicaltrials.gov: NCT01641822.

Cystic fibrosis (CF) lung infections caused by members of the Burkholderia cepacia complex, such as Burkholderia multivorans, are associated with high rates of mortality and morbidity. We performed a population genomics study of 111 B. multivorans sputum isolates from one CF patient through three stages of infection including an early incident isolate, deep sampling of a one-year period of chronic infection occurring weeks before a lung transplant, and deep sampling of a post-transplant infection. We reconstructed the evolutionary history of the population and used a lineage-controlled genome-wide association study (GWAS) approach to identify genetic variants associated with antibiotic resistance. We found the incident isolate was basally related to the rest of the strains and more susceptible to antibiotics from three classes (β-lactams, aminoglycosides, quinolones). The chronic infection isolates diversified into multiple, distinct genetic lineages and showed reduced antimicrobial susceptibility to the same antibiotics. The post-transplant reinfection isolates derived from the same source as the incident isolate and were genetically distinct from the chronic isolates. They also had a level of susceptibility in between that of the incident and chronic isolates. We identified numerous examples of potential parallel pathoadaptation, in which multiple mutations were found in the same locus or even codon. The set of parallel pathoadaptive loci was enriched for functions associated with virulence and resistance. Our GWAS analysis identified statistical associations between a polymorphism in the ampD locus with resistance to β-lactams, and polymorphisms in an araC transcriptional regulator and an outer membrane porin with resistance to both aminoglycosides and quinolones. Additionally, these three loci were independently mutated four, three and two times, respectively, providing further support for parallel pathoadaptation. Finally, we identified a minimum of 14 recombination events, and observed that loci carrying putative parallel pathoadaptations and polymorphisms statistically associated with β-lactam resistance were over-represented in these recombinogenic regions., Author summary Cystic fibrosis (CF) is the most common lethal genetic disorder affecting individuals of European descent. Most CF patients die at a young age due to chronic lung infections. Among the organisms involved in these infections are bacteria from the Burkholderia cepacia complex (BCC), which are strongly associated with poor clinical prognosis. This study examines how the most prevalent BCC species among CF patients, B. multivorans, evolves within a single CF patient by studying the first B. multivorans isolate recovered from the patient, one hundred isolates recovered over a one year period during the chronic infection phase, and an additional ten isolates recovered after the reinfection of the transplanted lungs. We found that B. multivorans diversify phenotypically and genetically within the CF lung over the course of the infection, and evolves into a complex population during the chronic infection phase. We found that isolates collected from the post-transplant reinfection were more closely related to descendants of the original isolate rather than those recovered in the chronic infection. We identify genetic variants statistically associated with resistance to the antibiotics, and showed that some of these variants were found in regions that show patterns of recombination (genetic exchange) between strains. We also found that genes which were mutated multiple times during overall infection were more likely to be found in regions showing signals consistent with recombination. The presence of multiple independent mutations in a gene is a very strong signal that the gene helps bacteria adapt to their environment. Overall, this study provides insight into how pathogens adapt to the host during long-term infections, specific genes associated with antibiotic resistance, and the origin of new and recurrent infections.

Objective Determining the mechanisms that modulate β-lactam resistance in clinical Pseudomonas aeruginosa (P. aeruginosa) isolates can be challenging, as the molecular profiles identified in mutation-based or expression-based resistance determinant screens may not correlate with in vitro phenotypes. One of the lesser studied resistance mechanisms in P. aeruginosa is the modification of penicillin-binding protein 3 (pbpB/ftsI). This study reported that nonsynonymous polymorphisms within pbpB frequently occur among β-lactam resistant sputum isolates, and are associated with unique antibiotic susceptibility patterns. Methods Longitudinally collected isolates (n = 126) from cystic fibrosis (CF) patients with or without recent β-lactam therapy or of non-clinical origin were tested for susceptibility to six β-lactams (aztreonam, ceftazidime, cefsulodin, cefepime, meropenem, and piperacillin). Known β-lactam resistance mechanisms were characterised by polymerase chain reaction (PCR)-based methods, and polymorphisms in the transpeptidase-encoding domain of pbpB identified by sequencing. Results Twelve nonsynonymous polymorphisms were detected among 86 isolates (67%) from five CF patients with a history of β-lactam therapy, compared with one polymorphism in 30 (3.3%) from three patients who had not received β-lactam treatments. No nonsynonymous polymorphisms were found in ten environmental isolates. Multiple pbpB alleles, often with different combinations of polymorphisms, were detected within the population of strains from each CF patient for up to 2.6 years. Traditional patterns of ampC or mexA de-repression reduced expression of oprD or the presence of extended-spectrum β-lactamases were not observed in resistant isolates with nonsynonymous polymorphisms in pbpB. Conclusion This study's findings suggest that pbpB is a common adaptive target, and may contribute to the development of β-lactam resistance in P. aeruginosa.

Cystic fibrosis (CF) lung infections caused by members of the Burkholderia cepacia complex, such as Burkholderia multivorans, are associated with high rates of mortality and morbidity. We performed a population genomic study of 111 B. multivorans sputum isolates from a single CF patient through three stages of infection including the initial incident infection, deep sampling of a one-year period of chronic infection, and deep sampling of a post-transplant recolonization. We reconstructed the evolutionary history of the population and used a lineage-controlled genome-wide association study (GWAS) approach to identify genetic variants associated with antibiotic resistance. We found that the incident isolate was more susceptible to agents from three antimicrobial classes (β-lactams, aminoglycosides, quinolones), while the chronic isolates diversified into distinct genetic lineages with reduced antimicrobial susceptibility to the same agents. The post-transplant reinfection isolates displayed genetic and phenotypic signatures that were distinct from sputum isolates from all CF lung specimens. There were numerous examples of parallel pathoadaptation, in which individual loci, or even the same codon, were independently mutated multiple times. This set of loci was enriched for functions associated with virulence and resistance. Our GWAS approach identified one variant in the ampD locus (which was independently mutated four times in our dataset) associated with resistance to β-lactams, and two non-synonymous polymorphisms associated with resistance to both aminoglycosides and quinolones, affecting an araC family transcriptional regulator, which was independently mutated three times, and an outer member porin, which was independently mutated twice. We also performed recombination analysis and identified a minimum of 14 recombination events. Parallel pathoadaptive loci and polymorphisms associated with β-lactam resistance were over-represented in these recombinogenic regions. This study illustrates the power of deep, longitudinal sampling coupled with evolutionary and lineage-corrected GWAS analyses to reveal how pathogens adapt to their hosts.

Physician performance feedback (PPF) can help physicians gain insight into their practice, to identify areas for improvement, and to implement changes to improve care. There is increasing interest in the use of PPF in Canada. However, little is known about the different types of PPF methods and whether PPF can lead to improved physician performance and patient outcomes. We provide a primer for healthcare leaders interested in doing PPF by reviewing common PPF methods. We then describe our institution's experience with physician multi-source feedback and provide strategies to conduct meaningful PPF.

Over 90% of cystic fibrosis (CF) patients die due to chronic lung infections leading to respiratory failure. The decline in CF lung function is greatly accelerated by intermittent and progressively severe acute pulmonary exacerbations (PEs). Despite their clinical impact, surprisingly few microbiological signals associated with PEs have been identified. Here we introduce an unsupervised, systems-oriented approach to identify key members of the microbiota. We used two CF sputum microbiome data sets that were longitudinally collected through periods spanning baseline health and PEs. Key taxa were defined based on three strategies: overall relative abundance, prevalence, and co-occurrence network interconnectedness. We measured the association between changes in the abundance of the key taxa and changes in patient clinical status over time via change-point detection, and found that taxa with the highest level of network interconnectedness tracked changes in patient health significantly better than taxa with the highest abundance or prevalence. We also cross-sectionally stratified all samples into the clinical states and identified key taxa associated with each state. We found that network interconnectedness most strongly delineated the taxa among clinical states, and that anaerobic bacteria were over-represented during PEs. Many of these anaerobes are oropharyngeal bacteria that have been previously isolated from the respiratory tract, and/or have been studied for their role in CF. The observed shift in community structure, and the association of anaerobic taxa and PEs lends further support to the growing consensus that anoxic conditions and the subsequent growth of anaerobic microbes are important predictors of PEs., Cystic Fibrosis: Oxygen, microbes and exacerbation Episodes of significant worsening of cystic fibrosis symptoms, known as pulmonary exacerbations (PEs), are associated with oxygen-deficient (anoxic) conditions and increased activity of ‘anaerobic’ bacteria, which thrive in the absence of oxygen. Researchers in Canada, led by David Guttman at the University of Toronto, compared genetic data on microbial populations in sputum samples collected during PEs and at times of better health. The study revealed a strong correlation between the activity and interactions among anaerobic bacteria and the onset of PEs. Investigating the significance of these changes in the lung environment and its microbial populations may help design treatment strategies to reduce the frequency of PEs and their potentially fatal consequences. The authors suggest that antibiotics that specifically target anaerobic bacteria may prove beneficial, as may hyperbaric oxygen therapy, which oxygenates the lung tissue.

Achromobacter species are increasingly identified in individuals with cystic fibrosis (CF), but the clinical outcomes in these patients remain poorly understood.We aimed to determine the association of Achromobacter infection on clinical outcomes in pediatric and adult patients with CF.A cohort study of pediatric and adult patients with CF was conducted from 1997 to 2014 in Toronto, Ontario, Canada. Achromobacter spp. infection was categorized as no history of infection, intermittent infection, and chronic infection (two or more positive cultures in the preceding 12 months). Cox models were used to estimate risk of death or transplantation. Mixed-effects models were used to assess odds of pulmonary exacerbations and effect on lung function (FEVA total of 1,103 patients were followed-up over the course of 18 years; 88 patients (7.3%) had one or more culture for Achromobacter species. Chronic Achromobacter infection was associated with a greater risk of death or transplantation compared with in patients with no history of infection (adjusted hazard ratio, 2.03; 95% confidence interval, 1.05-3.95; P = 0.036). Pulmonary exacerbations were more common in patients with chronic infection, but after adjusting for confounding factors, the effect was no longer significant. The chronic group had lower FEVPatients with CF and chronic Achromobacter infection are at increased risk of death or transplantation.

BACKGROUND: Pulmonary exacerbations (PEx) are associated with acute loss of lung function that is often not recovered after treatment. We investigated lung function recovery following PEx for ivacaftor- and placebo-treated subjects. METHODS: Short- and long-term pulmonary function recovery data after PEx were summarized from a placebo-controlled trial in 161 cystic fibrosis patients≥12years old with the G551D-CFTR mutation (NCT00909532). Short-term recovery was measured 2 to 8weeks after treatment, and long-term recovery was determined at the end-of-study, both compared with baseline measured just prior to the PEx. RESULTS: Fewer patients receiving ivacaftor experienced a PEx than patients receiving placebo (33.7% vs. 56.4%; P=0.004) and had a lower adjusted incidence rate of PEx (0.589 vs. 1.382; P

Background Individuals with Burkholderia spp. infection have historically been excluded from efficacy trials of inhaled antibiotics, including aztreonam for inhalation solution (AZLI). Methods A double-blind, placebo-controlled, 24-week trial of continuous AZLI/placebo treatment was undertaken in individuals with cystic fibrosis (CF) and chronic Burkholderia spp. infection. All subjects also received usual medical care (determined by their physicians). Additional antibiotic use was not restricted. Results Baseline FEV 1 % predicted values ranged from 15.8% to 114.6%. No significant treatment differences (AZLI vs. placebo) were observed at week 24 for any endpoints, including FEV 1 % predicted, number of respiratory exacerbations requiring systemic/inhaled antibiotics, or hospitalizations. Continuous AZLI administration was well tolerated. Burkholderia spp. susceptibility to antibiotics commonly used in CF therapy showed little change. Conclusions 24-weeks of continuous AZLI treatment did not significantly improve lung function in CF subjects with chronic Burkholderia spp. infection. Non-study antibiotic use may have confounded any potential AZLI effects.

Background Biobanks are considered to be key infrastructures for research development and have generated a lot of debate about their ethical, legal and social implications (ELSI). While the focus has been on human genomic research, rapid advances in human microbiome research further complicate the debate. Discussion We draw on two cystic fibrosis biobanks in Toronto, Canada, to illustrate our points. The biobanks have been established to facilitate sample and data sharing for research into the link between disease progression and microbial dynamics in the lungs of pediatric and adult patients. We begin by providing an overview of some of the ELSI associated with human microbiome research, particularly on the implications for the broader society. We then discuss ethical considerations regarding the identifiability of samples biobanked for human microbiome research, and examine the issue of return of results and incidental findings. We argue that, for the purposes of research ethics oversight, human microbiome research samples should be treated with the same privacy considerations as human tissues samples. We also suggest that returning individual microbiome-related findings could provide a powerful clinical tool for care management, but highlight the need for a more grounded understanding of contextual factors that may be unique to human microbiome research. Conclusions We revisit the ELSI of biobanking and consider the impact that human microbiome research might have. Our discussion focuses on identifiability of human microbiome research samples, and return of research results and incidental findings for clinical management.

Pulmonary infections caused by Pseudomonas aeruginosa are a recalcitrant problem in cystic fibrosis (CF) patients. While the clinical implications and long-term evolutionary patterns of these infections are well studied, we know little about the short-term population dynamics that enable this pathogen to persist despite aggressive antimicrobial therapy. Here, we describe a short-term population genomic analysis of 233 P. aeruginosa isolates collected from 12 sputum specimens obtained over a 1-year period from a single patient. Whole-genome sequencing and antimicrobial susceptibility profiling identified the expansion of two clonal lineages. The first lineage originated from the coalescence of the entire sample less than 3 years before the end of the study and gave rise to a high-diversity ancestral population. The second expansion occurred 2 years later and gave rise to a derived population with a strong signal of positive selection. These events show characteristics consistent with recurrent selective sweeps. While we cannot identify the specific mutations responsible for the origins of the clonal lineages, we find that the majority of mutations occur in loci previously associated with virulence and resistance. Additionally, approximately one-third of all mutations occur in loci that are mutated multiple times, highlighting the importance of parallel pathoadaptation. One such locus is the gene encoding penicillin-binding protein 3, which received three independent mutations. Our functional analysis of these alleles shows that they provide differential fitness benefits dependent on the antibiotic under selection. These data reveal that bacterial populations can undergo extensive and dramatic changes that are not revealed by lower-resolution analyses., IMPORTANCE Pseudomonas aeruginosa is a bacterial opportunistic pathogen responsible for significant morbidity and mortality in cystic fibrosis (CF) patients. Once it has colonized the lung in CF, it is highly resilient and rarely eradicated. This study presents a deep sampling examination of the fine-scale evolutionary dynamics of P. aeruginosa in the lungs of a chronically infected CF patient. We show that diversity of P. aeruginosa is driven by recurrent clonal emergence and expansion within this patient and identify potential adaptive variants associated with these events. This high-resolution sequencing strategy thus reveals important intraspecies dynamics that explain a clinically important phenomenon not evident at a lower-resolution analysis of community structure.

The average age of lung transplant recipients is increasing, and the mix of recipient indications for transplantation is changing.To determine whether the health-related quality-of-life (HRQL) benefit of lung transplantation differs by recipient age and diagnosis.In this prospective cohort study, we obtained serial HRQL measurements in adults with advanced lung disease who subsequently underwent lung transplantation (2004-2012). HRQL assessments included the St. George's Respiratory Questionnaire, 36-Item Short-Form Health Survey (SF-36), EQ-5D, Standard Gamble, and Visual Analog Scale for current health. We used linear mixed effects models for associations between age or diagnosis and changes in HRQL with transplantation. To address potential survivorship bias, we fitted Markov models to the distribution of discrete post-transplant health states (HRQL better than pretransplant, not better, or dead) and estimated quality-adjusted life-years post-transplant.A total of 430 subjects were listed, 387 were transplanted, and 326 provided both pretransplant and post-transplant data. Transplantation conferred large improvements in all HRQL measures: St. George's change of -47 units (95% confidence interval, -48 to -44), 36-Item Short-Form Health Survey physical component summary score of 17.7 (16.5-18.9), EQ-5D of 0.27 (0.24-0.30), Standard Gamble of 0.48 (0.44-0.51), and Visual Analog of 44 (42-47). Age was not associated with meaningful differences in the HRQL benefits of transplantation. There was less HRQL benefit in interstitial lung disease than in cystic fibrosis.Lung transplantation confers large HRQL benefits, which vary by recipient diagnosis, but do not differ substantially in older recipients.

Chronic airway infections caused by Pseudomonas aeruginosa contribute to the progression of pulmonary disease in individuals with cystic fibrosis (CF). In the setting of CF, within-patient adaptation of a P. aeruginosa strain generates phenotypic diversity that can complicate microbiological analysis of patient samples. We investigated within- and between- sample diversity of 34 phenotypes among 235 P. aeruginosa isolates cultured from sputum samples collected from a single CF patient over the span of one year and assessed colony morphology as a screening tool for predicting phenotypes, including antimicrobial susceptibilities. We identified 15 distinct colony morphotypes that varied significantly in abundance both within and between sputum samples. Substantial within sample phenotypic heterogeneity was also noted in other phenotypes, with morphotypes being unreliable predictors of antimicrobial susceptibility and other phenotypes. Emergence of isolates with reduced susceptibility to β-lactams was observed during periods of clinical therapy with aztreonam. Our findings confirm that the P. aeruginosa population in chronic CF lung infections is highly dynamic and that intra-sample phenotypic diversity is underestimated if only one or few colonies are analyzed per sample.

Understanding the significance of bacterial species that colonize and persist in cystic fibrosis (CF) airways requires a detailed examination of bacterial community structure across a broad range of age and disease stage. We used 16S ribosomal RNA sequencing to characterize the lung microbiota in 269 CF patients spanning a 60 year age range, including 76 pediatric samples from patients of age 4–17 and a broad cross-section of disease status to identify features of bacterial community structure and their relationship to disease stage and age. The CF lung microbiota shows significant inter-individual variability in community structure, composition and diversity. The core microbiota consists of five genera - Streptococcus, Prevotella, Rothia, Veillonella and Actinomyces. CF-associated pathogens such as Pseudomonas, Burkholderia, Stenotrophomonas and Achromobacter are less prevalent than core genera, but have a strong tendency to dominate the bacterial community when present. Community diversity and lung function are greatest in patients less than 10 years of age and lower in older age groups, plateauing at approximately age 25. Lower community diversity correlates with worse lung function in a multivariate regression model. Infection by Pseudomonas correlates with age-associated trends in community diversity and lung function.

Cystic fibrosis (CF) related diabetes mellitus (DM) occurs in 15% of adult pancreatic insufficient CF patients. Lung transplantation is a treatment option for end-stage CF. We hypothesized that the prevalence of DM increases after lung transplantation. The study population included adult patients undergoing lung transplantation from March 1988 to March 2002 for end-stage CF at the University of Toronto. Demographic data, exocrine pancreatic function, presence of DM before and after transplant, as well as timing of its development after transplant were collected. Eighty-six patients met the study criteria; 77 of 86 (89.5%) of patients were pancreatic insufficient and were further analyzed. Median follow-up post-transplant was 3.3 yr (interquartile range: 1.2-7.2). Their mean age was 29.7 +/- 8.1 yr and 46 of 77 (59.7%) were male. The prevalence of DM increased from 22 of 77 (28.6%) before transplant to 38 of 77 (49.4%) after transplant (p = 0.008). The median time of DM development after transplant was 80 d (range: 13-4352). Sixteen of 55 (29.1%) of pancreatic insufficient patients who were non-diabetic prior to transplant, developed DM after transplant. DM is common in CF patients undergoing lung transplantation and the prevalence increases after transplant.

Study objective To determine the repeatability of measurements of FEV 1 in adults with lung disease due to cystic fibrosis (CF). Design Single cohort study nested within a randomized controlled trial. Setting Adult CF of a university teaching hospital. Subjects were participants in a randomized trial of an experimental mucolytic drug. Patients Twenty-one adults (mean age, 27.5 ± 9.2 years [± SD]) with CF and mild-to-moderate airflow obstruction (FEV 1 70 ± 15% predicted). Patients were in clinically stable condition prior to and during the study. Interventions Repeated FEV 1 measurements were obtained at specific times of the day for 9 consecutive days, for a total of 31 measurements from each subject. Statistical measures of repeatability were calculated. Variation over the course of 1 day and variation from 1 day to the next were examined separately. Measurements and results For day-to-day FEV 1 measurements, the within-subject SD was 0.145 L (4.5% of predicted), indicating greater variation compared to values previously established in normal subjects. The coefficient of repeatability indicated that day-to-day measurements could differ by as much as 13% of predicted in the absence of clinical change. For measurements within a single day, variation was not observed to be greater than normal. Conclusions In adults with CF, day-to-day variation in FEV 1 measurements is greater than normal and similar to that seen in other obstructive lung diseases. Changes in FEV 1 over time in adults with CF can likely be interpreted using the same criteria that apply to asthma or COPD.

Objective To assess the effect of pregnancy on pulmonary function and survival in women with cystic fibrosis (CF) and to assess the fetal outcome. Design Cohort study. The data analyzed were collected from the Toronto CF database, chart review, and patient questionnaire. Setting Tertiary-care center. Patients All women with CF who, at the time of diagnosis or pregnancy, attended the Toronto Cystic Fibrosis Clinics between 1961 and 1998. Results From 1963 to 1998, there were 92 pregnancies in 54 women. There were 11 miscarriages and 7 therapeutic abortions. Forty-nine women gave birth to 74 children. The mean follow-up time was 11 ± 8 years. One patient was lost to follow-up shortly after delivery, and one was lost after 12 years. The overall mortality rate was 19% (9 of 48 patients). Absence of Burkholderia cepacia (p 1 > 50% predicted (p = 0.03) were associated with better survival rates. When adjusted for the same parameters, pregnancy did not affect survival compared to the entire adult female CF population. The decline in FEV 1 was comparable to that in the total CF population. Three women had diabetes mellitus, and seven developed gestational diabetes. There were six preterm infants and one neonatal death. CF was diagnosed in two children. Conclusions The maternal and fetal outcome is good for most women with CF. Risk factors for mortality are similar to those for the nonpregnant CF population. Pregnancies should be planned so that there is opportunity for counseling and optimization of the medical condition. Good communication between the CF team and the obstetrician is important.

Study objectives To determine (1) the prevalence of pulmonary hypertension and cardiac dysfunction in adult cystic fibrosis (CF) patients with severe lung disease, (2) the relationship between these cardiovascular abnormalities and hypoxemia, and (3) the impact of subclinical pulmonary hypertension on survival. Design Single-blind, cross-sectional study. Setting Ambulatory clinic of the Adult CF program at a tertiary-level hospital. Patients Clinically stable patients with severe lung disease (FEV1 Measurements and results Eighteen patients with severe lung disease (FEV1 28 ± 7% of predicted normal value) were initially studied. Each patient had overnight polysomnography, pulmonary function tests, and Doppler echocardiography. Arterial oxygen saturation (Sao2) was reduced during wakefulness (87.1 ± 6.1%) and fell during sleep (84.0 ± 6.6%) while transcutaneous Pco2 was normal during wakefulness (41.1 ± 6.9 mm Hg) and increased during sleep (46.6 ± 4.7 mm Hg). Left ventricular size, systolic function, and diastolic function were normal except in one patient who had had a previous silent myocardial infarction due to coronary artery disease. Qualitative assessment of right ventricular function was normal in all patients. Pulmonary artery systolic pressure (PASP) was increased (> 35 mm Hg) in seven patients without clinical evidence of cor pulmonale. Regression analysis was performed by combining these data with data from an additional 15 CF patients with moderately severe lung disease (FEV1 56.3 ± 8.9% predicted normal) who were recruited to a modified study protocol that included overnight oximetry, pulmonary function tests, and Doppler echocardiography. None of these patients had evidence of hypoxemia and only three had mild elevation of PASP (36, 37, and 39 mm Hg). Linear regression analysis revealed that PASP was significantly correlated with FEV1(r = −0.44; p=0.013), and Sao2 during wakefulness (r = −0.60; p=0.0003), during sleep (r = −0.56; p=0.0008), and after 6 min of exercise (r = −0.75; p Conclusions In adult CF patients with severe stable lung disease, left and right ventricular function is well maintained in the absence of significant coronary artery disease; pulmonary hypertension develops in a significant proportion of patients and is strongly correlated with oxygen status, independent of lung function; and subclinical pulmonary hypertension is associated with an increased mortality.

We investigated the utility of PCR to detect Burkholderia cepacia directly in sputum samples at two cystic fibrosis (CF) centers serving children and adults. Following liquefaction of the sputa by using N -acetyl- l -cysteine, DNA was isolated and analyzed by PCRs with three different primer pairs directed toward bacterial rRNA loci. Two primer pairs were putatively specific for B. cepacia . The other pair, which universally amplifies a band from all bacteria, served as a control. Sputum samples were obtained from 219 patients and analyzed independently by culture and by PCR to detect B. cepacia . The analyses were performed blinded with respect to each other. The results of the PCR with sputa demonstrated that the primers directed to the 16S loci demonstrated approximately 95% concordance with culture results and were more specific than those amplifying the 16S to 23S spacer region. In addition, the 16S primer pair putatively identified B. cepacia in seven patients whose sputa were culture negative at this time. Of these culture-negative patients, five had sputum samples that were culture positive for B. cepacia either prior or subsequent to this study. The results of this study indicate the utility of PCR as a diagnostic method for the rapid identification of B. cepacia in sputum samples of CF patients. We anticipate that improvements in our taxonomic understanding may allow the design of more specific primers for detection of each species of the B. cepacia complex in sputum samples.

Burkholderia (Pseudomonas) cepaciahas been cultured from bronchial secretions of patients with cystic fibrosis with varying frequency. The overall prevalence in patients over 18 years old at the authors’ centre in Toronto is 51%. Extrapulmonary infections are rare in nonimmunosuppressed patients with cystic fibrosis andB cepaciahas been the causative organism in only three previous reports. Two patients with cystic fibrosis are described who developed hepatic abscesses clue toB cepacia. One patient is the sibling of two brothers in whom this complication was previously described.

The characterization of bacterial communities using DNA sequencing has revolutionized our ability to study microbes in nature and discover the ways in which microbial communities affect ecosystem functioning and human health. Here we describe Serial Illumina Sequencing (SI-Seq): a method for deep sequencing of the bacterial 16S rRNA gene using next-generation sequencing technology. SI-Seq serially sequences portions of the V5, V6 and V7 hypervariable regions from barcoded 16S rRNA amplicons using an Illumina short-read genome analyzer. SI-Seq obtains taxonomic resolution similar to 454 pyrosequencing for a fraction of the cost, and can produce hundreds of thousands of reads per sample even with very high multiplexing. We validated SI-Seq using single species and mock community controls, and via a comparison to cystic fibrosis lung microbiota sequenced using 454 FLX Titanium. Our control runs show that SI-Seq has a dynamic range of at least five orders of magnitude, can classify >96% of sequences to the genus level, and performs just as well as 454 and paired-end Illumina methods in estimation of standard microbial ecology diversity measurements. We illustrate the utility of SI-Seq in a pilot sample of central airway secretion samples from cystic fibrosis patients.

Background. Recurrent bacterial infections play a key role in the pathogenesis of bronchiectasis, but conventional microbiologic methods may fail to identify pathogens in many cases. We characterized and compared the pulmonary bacterial communities of cystic fibrosis (CF) and non-CF bronchiectasis patients using a culture-independent molecular approach.Methods. Bacterial 16S rRNA gene libraries were constructed from lung tissue of 10 non-CF bronchiectasis and 21 CF patients, followed by DNA sequencing of isolates from each library. Community characteristics were analyzed and compared between the two groups.Results. A wide range of bacterial diversity was detected in both groups, with between 1 and 21 bacterial taxa found in each patient.Pseudomonaswas the most common genus in both groups, comprising 49% of sequences detected and dominating numerically in 13 patients. AlthoughPseudomonasappeared to be dominant more often in CF patients than in non-CF patients, analysis of entire bacterial communities did not identify significant differences between these two groups.Conclusions. Our data indicate significant diversity in the pulmonary bacterial community of both CF and non-CF bronchiectasis patients and suggest that this community is similar in surgically resected lungs of CF and non-CF bronchiectasis patients.

Rationale There is no adequate explanation for gender-based differences in rates of mortality and of deterioration in pulmonary function in cystic fibrosis (CF) patients. One potential explanation is that gender hormones (sex steroids) may modulate the severity of CF lung disease, the principal cause of mortality in CF, by altering respiratory transepithelial ion transport. Objective To determine whether respiratory epithelial ion transport varied during the menstrual cycle of CF females. Methods The nasal transepithelial electrical potential difference (NPD) was determined as a measure of ion transport across human respiratory epithelium, coincident with measurements of endogenous serum hormone levels in the luteal and follicular phases of the menstrual cycle in CF females aged 16–22 years. Results The component of the NPD that is insensitive to the Na+ transport blocker amiloride, but not the amiloride-sensitive component, changed in association with endogenous, menstrual cycle-induced changes in serum levels of progesterone and estrogen (P = 0.02, n = 7, paired t-test). Measurements using Cl− free perfusates suggested that the changes are not a result of Cl− conductance. Conclusions Our results suggest that in CF respiratory epithelium amiloride-insensitive, but not amiloride-sensitive, ion transport is altered by female gender hormones in vivo. We speculate that amiloride-insensitive ion transport may contribute to the regulation of human airway surface fluid. Pediatr Pulmonol. 2007; 42:519–524. © 2007 Wiley-Liss, Inc.

Cystic fibrosis (CF) related diabetes mellitus (DM) occurs in 15% of adult pancreatic insufficient CF patients. Lung transplantation is a treatment option for end-stage CF. We hypothesized that the prevalence of DM increases after lung transplantation. The study population included adult patients undergoing lung transplantation from March 1988 to March 2002 for end-stage CF at the University of Toronto. Demographic data, exocrine pancreatic function, presence of DM before and after transplant, as well as timing of its development after transplant were collected. Eighty-six patients met the study criteria; 77 of 86 (89.5%) of patients were pancreatic insufficient and were further analyzed. Median follow-up post-transplant was 3.3 yr (interquartile range: 1.2-7.2). Their mean age was 29.7 +/- 8.1 yr and 46 of 77 (59.7%) were male. The prevalence of DM increased from 22 of 77 (28.6%) before transplant to 38 of 77 (49.4%) after transplant (p = 0.008). The median time of DM development after transplant was 80 d (range: 13-4352). Sixteen of 55 (29.1%) of pancreatic insufficient patients who were non-diabetic prior to transplant, developed DM after transplant. DM is common in CF patients undergoing lung transplantation and the prevalence increases after transplant.

Objective: To define the clinical characteristics and diagnostic parameters of patients with cystic fibrosis (CF) diagnosed in adulthood. Design: Retrospective cohort study. Setting: Tertiary care center. Patients and methods: All patients with a diagnosis of CF made at the Toronto CF Clinics between 1960 and June 2001. Data were collected prospectively and analyzed retrospectively. Results: There were 73 of 1,051 patients (7%) with CF diagnosed in adulthood. Over time, an increasing number and proportion of patients received a diagnosis in adulthood: 27 patients (3%) before 1990, compared to 46 patients (18%) after 1990 (p < 0.001). The mean sweat chloride level was lower for those with CF diagnosed as adults, compared to those with a diagnosis as children (75 26 mmol/L and 100 19 mmol/L, respectively; p < 0.001) [mean SD], and adults were more likely to have pancreatic sufficiency (PS) than children (73% vs 13%, respectively; p < 0.0001). In 46 adults who received a diagnosis since 1990, the reason for the initial sweat test was pancreatitis (2 patients, 4%), pulmonary symptoms (18 patients, 39%), pulmonary and GI symptoms (10 patients, 22%), infertility (12 patients, 26%), and genetic screening (4 patients, 9%). Other manifestations were biliary cirrhosis (one patient) and diabetes mellitus (four patients, 9%). The diagnosis could be confirmed by sweat test alone in 30 of 46 patients (65%), by mutation analysis alone in 15 patients (33%), and by a combination in 31 patients (67%). Nasal potential difference (PD) measurements alone confirmed the diagnosis in the remaining 15 patients (33%). Conclusion: Patients with CF presenting in adulthood often have PS, inconclusive sweat test results, and a high prevalence of mutations that are not commonly seen in CF diagnosed in childhood. Although most patients have lung disease of variable degrees, single-organ manifestations such as congenital bilateral absence of the vas deferens and pancreatitis are seen. Repeated sweat tests and extensive mutation analysis are often required. Nasal PD may aid the diagnosis, but has not been standardized for clinical diagnosis. (CHEST 2004; 126:1215–1224)

In cystic fibrosis (CF), airway disease begins early in life. Bacteria and elevated levels of neutrophils and inflammatory mediators have been detected in bronchoalveolar lavage (BAL) fluid from infants with CF. Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) are common in men with congenital bilateral absence of the vas deferens (CBAVD) and it has been suggested that this syndrome represents a mild form of CF. We hypothesized that men with CBAVD also have subclinical pulmonary disease. Bronchoscopy with BAL, viral and quantitative bacterial cultures, and analyses of total and differential cell count, cytokines, and free neutrophil elastase was performed in eight men with CBAVD, who had mutations in the CFTR and intermediate or elevated sweat chloride levels, and in four healthy control subjects. There was light growth of Staphylococcus aureus in one of eight men with CBAVD, and small numbers of opportunistic gram-negative bacteria in six of eight men with CBAVD and in one control subject. BAL cell counts and neutrophil elastase were within the normal range. Interleukin-8 and tumor necrosis factor-alpha levels were higher for men with CBAVD than for control subjects. These data suggest that mutations in the CFTR in men with CBAVD, in addition to causing infertility, lead to subclinical bacterial pulmonary infection and inflammation consistent with mild CF.

Study Objective To ascertain the incidence, types, morbidity, and mortality of infectious episodes in isolated lung transplant recipients. Design Retrospective chart review of patients who have undergone transplants over a six-year period in one institution. Patients Twenty-three single and 17 double lung transplants followed up between 2 and 68 months. Results Fifty-one episodes of infection occurred in the group with a slight predominance in the double lung transplants. The 32 episodes of bacterial infection constituted the largest group of infection and more than half of these were pneumonias. Organisms identified were predominantly Gram negative. While bacterial processes made up the bulk of infections, fatalities were rare. Viral and fungal infections were less common, but more often fatal. Of six cases of viral pneumonitis, two were fatal; two of five cases of invasive fungal infection were also fatal. Overall, six patients died of infection. Conclusion Our findings support previous reports from heart-lung centers documenting a high rate of infectious complications, particularly pneumonia, in recipients of lung grafts. In our experience, bacterial infections are the most common (two of three infections), but have the lowest mortality. Efforts should be directed toward establishing effective prophylaxis programs and early detection of infection.